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Testing AAVrg-hSyn-mCherry in the Putamen of Rhesus Macaques

Addgene #114472-AAVrg

Data Submitted By
Adriana Galvan and Johnson Agniswamy
Submission Date
August 15, 2023
Publication Date
August 24, 2023 (modified August 24, 2023)
DOI
https://doi.org/10.57733/addgene.pwsycm
Funding
Aligning Science Across Parkinson's ASAP-020572 through the Michael J. Fox Foundation for Parkinson’s Research (MJFF) ( Wichmann, PI, Galvan Co-I); P50NS123103 (NIH/NINDS) (Wichmann, PI, Galvan Co-I); P51-OD011132 (Emory Primate Center base grant); ASAP-020572; P50NS123103; P51-OD011132
Abstract
Two rhesus macaque animals were tested for AAVrg-hSyn-mCherry in the putamen. We observed abundant mCherry-positive neurons in expected targets (i.e., motor cortical regions known to project to the putamen).
Citation
Testing AAVrg-hSyn-mCherry in the Putamen of Rhesus Macaques. Galvan A, Agniswamy J 2023. Addgene Report, https://doi.org/10.57733/addgene.pwsycm
Injection of AAVrg-hSyn-mCherry in the putamen of rhesus monkey. (A) Low magnification view of a coronal section of a rhesus macaque that received AAVrg-hSyn-mCherry in the putamen. B to E) Higher magnification images of areas shown in rectangles in A. Abundant mCherry positive cells were found in M1 and SMA. In the external globus pallidus (GPe), we found a combination of mCherry-positive putative terminals and cell bodies (arrow), while in GPi, we found mCherry-positive putative terminals. The labeling of terminals in GPe and GPi indicates anterograde transport from the site of injection (putamen)
Caption: Injection of AAVrg-hSyn-mCherry in the putamen of rhesus monkey. (A) Low magnification view of a coronal section of a rhesus macaque that received AAVrg-hSyn-mCherry in the putamen. B to E) Higher magnification images of areas shown in rectangles in A. Abundant mCherry positive cells were found in M1 and SMA. In the external globus pallidus (GPe), we found a combination of mCherry-positive putative terminals and cell bodies (arrow), while in GPi, we found mCherry-positive putative terminals. The labeling of terminals in GPe and GPi indicates anterograde transport from the site of injection (putamen)

This report is made available under the Creative Commons Attribution 4.0 International License.

Vector Used
pAAV-hSyn-mCherry
Addgene #114472-AAVrg

Virus & Injection

Virus

Virus Name
pAAV-hSyn-mCherry
Serotype
AAVrg
Source
Addgene
Catalog Number
114472-AAVrg
Virus Lot Number
v111416
Promoter
Synapsin
Cargo Type
Control
Injection Titer / Dose
2.00 E+13 vg/ml
Injection Volume
2 tracks of 2 µl each (total 4 µl)
Injection Rate
0.5 µl/min
Injector Material
Hamilton micro syringe
Tip Diameter
30
Injection Time
For putamen injections, the needle was left in place for 10 minutes before withdrawal.
Injection Site / Route
Post-commissural putamen. Neuronames ID: 230.
Additional Details
Targeting and trajectory planning occurred through presurgical magnetic resonance imaging (MRI). Two injections tracks were done, in each we deposited 2 µl of the viral vector (total 4 μL), at an infusion rate of 0.5 μL/min using a Hamilton microsyringe (Reno, NE) coupled to a stereotax-mounted automated injector pump system.
View in Addgene Catalog

Other Details

Species
Non-Human Primate
Strain or Cell Line
Rhesus Macaque
Source
Emory National Primate Research Center
Other Information
2 females
Age at Injection
5 years, 2 months; 5 y 6 months
Sex
Female
Time After Injection
6 and 8 months
Detection Method
IHC
Assay & Results
Two animals were injected in putamen. We used antibodies against mCherry to examine expression. The brain sections were pretreated (10% normal goat serum, 1% bovine serum albumin, and 0.3% Triton X-100) and then incubated in the antibody solution for 24 h. The sections were then incubated in the biotinylated antibody for 90 min. Afterward, the sections were placed in 0.025% 3-3’-diaminobenzidine tetrachloride, 0.01 M Imidazole, and 0.006% H2O2 for 10 min. The sections were mounted on slides, covered, and digitized with an Aperio Scanscope CS system.
Target Cell Transduction
We observed abundant mCherry-positive neurons in expected targets (i.e., motor cortical regions known to project to the putamen)